Yuan Ze-Chun, Zaheer Rahat, Finan Turlough M
Center for Environmental Genomics, Department of Biology, McMaster University, Hamilton, Ontario, Canada L8S 4K1.
J Bacteriol. 2006 Feb;188(3):1089-102. doi: 10.1128/JB.188.3.1089-1102.2006.
The properties and regulation of the pstSCAB-encoded Pi uptake system from the alfalfa symbiont Sinorhizobium meliloti are reported. We present evidence that the pstSCAB genes and the regulatory phoUB genes are transcribed from a single promoter that contains two PhoB binding sites and that transcription requires PhoB. S. meliloti strain 1021 (Rm1021) and its derivatives were found to carry a C deletion frameshift mutation in the pstC gene (designated pstC1021) that severely impairs activity of the PstSCAB Pi transport system. This mutation is absent in RCR2011, the parent of Rm1021. Correction of the pstC1021 mutation in Rm1021 by site-directed mutagenesis revealed that PstSCAB is a Pi-specific, high-affinity (Km, 0.2 microM), high-velocity (Vmax, 70 nmol/min/mg protein) transport system. The pstC1021 allele was shown to generate a partial pho regulon constitutive phenotype, in which transcription is activated by PhoB even under Pi-excess conditions that render PhoB inactive in a wild-type background. The previously reported symbiotic Fix- phenotype of phoCDET mutants was found to be dependent on the pstC1021 mutation, as Rm1021 phoCDET mutants formed small white nodules on alfalfa that failed to reduce N2, whereas phoCDET mutant strains with a corrected pstC allele (RmP110) formed pink nodules on alfalfa that fixed N2 like the wild type. Alfalfa root nodules formed by the wild-type RCR2011 strain expressed the low-affinity orfA-pit-encoded Pi uptake system and neither the pstSCAB genes nor the phoCDET genes. Thus, metabolism of alfalfa nodule bacteroids is not Pi limited.
本文报道了苜蓿共生菌苜蓿中华根瘤菌（Sinorhizobium meliloti）中由pstSCAB编码的磷摄取系统的特性和调控。我们提供的证据表明，pstSCAB基因和调控性phoUB基因是从一个包含两个PhoB结合位点的单一启动子转录而来，且转录需要PhoB。苜蓿中华根瘤菌1021菌株（Rm1021）及其衍生物在pstC基因中存在一个C缺失移码突变（命名为pstC1021），该突变严重损害了PstSCAB磷转运系统的活性。Rm1021的亲本RCR2011中不存在此突变。通过定点诱变纠正Rm1021中的pstC1021突变后发现，PstSCAB是一种磷特异性、高亲和力（Km，0.2微摩尔）、高速度（Vmax，70纳摩尔/分钟/毫克蛋白质）的转运系统。pstC1021等位基因表现出部分pho调控子组成型表型，即即使在使PhoB在野生型背景中失活的磷过量条件下，转录也会被PhoB激活。之前报道的phoCDET突变体的共生固氮缺陷型表型被发现依赖于pstC1021突变，因为Rm1021 phoCDET突变体在苜蓿上形成了不能还原N2的小白结节，而具有校正pstC等位基因的phoCDET突变体菌株（RmP110）在苜蓿上形成了能像野生型一样固定N2的粉红色结节。野生型RCR2011菌株形成的苜蓿根瘤表达低亲和力的orfA - pit编码的磷摄取系统，而不表达pstSCAB基因或phoCDET基因。因此，苜蓿根瘤类菌体的代谢不受磷限制。
